Mechanisms of Antimicrobial, Cytolytic, and Cell-Penetrating Peptides: From Kinetics to Thermodynamics

Abstract

The mechanisms of six different antimicrobial, cytolytic, and cell-penetrating peptides, including some of their variants, are discussed and compared. The specificity of these polypeptides varies, but they all form amphipathic α-helices when bound to membranes, and there are no striking differences in their sequences. We have examined the thermodynamics and kinetics of their interaction with phospholipid vesicles, namely binding and peptide-induced dye efflux. The thermodynamics of binding calculated using the Wimley-White interfacial hydrophobicity scale are in good agreement with the values derived from experiment. The generally accepted view that binding affinity determines functional specificity is also supported by experiment in model membranes. We now propose the hypothesis that it is the thermodynamics of peptide insertion into the membrane, from a surface-bound state, that determines the mechanism.During the past three decades, a vast number of antimicrobial peptides (1,2) and other related cytolytic peptides (3) have been discovered and their mechanisms examined. More recently, several cell-penetrating peptides have been described, which allow for transport of large molecules, such as proteins or DNA fragments, into cells (4-8). Perhaps surprisingly, many of these antimicrobial, cytolytic, and cell-penetrating peptides fall into the same structural class: they form an amphipathic α-helix of some 14-40 residues, when bound to a membrane surface. Yet, they show remarkable specificity regarding the target membrane or organism. What has befuddled researchers for a long time is the absence of a correlation between sequence and function or mechanism. The only element that appears to separate antimicrobial from cytolytic peptides is that antimicrobials are usually cationic. This provides a simple explanation for their specificity because cationic peptides should bind better to the anionic membranes of most bacteria than to the neutral membranes of eukaryotic cells (9).We now critically review results obtained over the past several years on a set of representative antimicrobial, cytolytic, and cell-penetrating peptides. The interactions of these peptides with model membranes were all studied with the same methods and under similar conditions. Experiments using small unilamellar vesicles (SUV) 1 were common in the past but we purposely exclude them because of the strained nature of those vesicles, concentrating instead on studies that use unstrained vesicles, such as large (LUV) or giant unilamellar vesicles (GUV). The results are, therefore, directly comparable. On the basis of a quantitative analysis of the kinetics and thermodynamics of these interactions, we propose the hypothesis that the peptide sequence only specifies the mechanism indirectly, through the thermodynamics of peptide insertion into the bilayer medium from the surface-bound state. This would explain the lack of direct correlation between sequence and mechanism. † This work was supported by National Institutes of Health Grant GM072507....

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